• Journal of Microbiology and Biotechnology
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• v.26 no.11
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• pp.1891-1907
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• 2016

Yeasts that are present in marine environments have evolved to survive hostile environments that are characterized by high exogenous salt content, high concentrations of inhibitory compounds, and low soluble carbon and nitrogen levels. Therefore, yeasts isolated from marine environments could have interesting characteristics for industrial applications. However, the application of marine yeast in research or industry is currently very limited owing to the lack of a suitable isolation method. Current methods for isolation suffer from fungal interference and/or low number of yeast isolates. In this paper, an efficient and non-laborious isolation method has been developed and successfully isolated large numbers of yeasts without bacterial or fungal growth. The new method includes a three-cycle enrichment step followed by an isolation step and a confirmation step. Using this method, 116 marine yeast strains were isolated from 14 marine samples collected in the UK, Egypt, and the USA. These strains were further evaluated for the utilization of fermentable sugars (glucose, xylose, mannitol, and galactose) using a phenotypic microarray assay. Seventeen strains with higher sugar utilization capacity than the reference terrestrial yeast Saccharomyces cerevisiae NCYC 2592 were selected for identification by sequencing of the ITS and D1/D2 domains. These strains belonged to six species: S. cerevisiae, Candida tropicalis, Candida viswanathii, Wickerhamomyces anomalus, Candida glabrata, and Pichia kudriavzevii. The ability of these strains for improved sugar utilization using seawater-based media was confirmed and, therefore, they could potentially be utilized in fermentations using marine biomass in seawater media, particularly for the production of bioethanol and other biochemical products.

• Korean Journal of Microbiology
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• v.42 no.4
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• pp.319-325
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• 2006

[ $\beta$ ]-Glucans (AG) were prepared from Agaricus blazei cultured in the medium fortified with the roots of Pueraria spp. by repeated extraction with hot water, gel filtration chromatography and DEAE ion exchange chromatography. Oligosaccharides (AO) were derived from the hydrolysis of AG by an endo-$\beta$-(1$\rightarrow$6)-glucanase from Bacillus megaterium. The anti-HT-29 human colon cancer activity of AG or AO was investigated using MTT assay, apoptosis assay, cell cycle analysis, and cDNA microairay. AG and AO both inhibited proliferation and growth of HT-29 cells, and stimulated apoptosis of the cells in a dose-dependent manner. In cell cycle analysis, treating HT-29 cells with AG or AO resulted in the increase of cells in the G0 (sub-G1) and G1 phase. Especially, AO was more effective in inducing G0/G1 cell cycle arrest than AG. To screen the genes involved in the increase of apoptosis, the gene expression profile of the HT-29 cells treated with AO was examined by cDNA microarray. While several genes involved in cell cycle progression (CCND2 and CDK2) were down-regulated, many genes involved in apoptosis (TNFSF9, TNFRSF9, FADD, CASP8, BAD, CRADD, CASP9 etc), cell cycle inhibitor (CDKN2A), immune response (IL6, IL18, IL6R etc), and tumor suppressor (CEACAM1, TP53BP2, IRF1, and PHB) were up-regulated. These results suggest that AO could inhibit the proliferation and growth of HT-29 cells by G0/G1 cell cycle arrest and induction of apoptosis.

• Biomolecules & Therapeutics
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• v.15 no.4
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• pp.252-260
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• 2007

As it is needed to assay possible feasibility of extrapolation between in vivo and in vitro systems and to develop a new in vitro method for toxicity testing, we investigated global gene expression from both animal and cell line treated with thioacetamide (TAA) and compared between in vivo and in vitro genomic profiles. For in vivo study, mice were orally treated with TAA and sacrificed at 6 and 24 h. For in vitro study, TAA was administered to a mouse hepatic cell line, BNL CL.2 and sampling was carried out at 6 and 24 h. Hepatotoxicity was assessed by analyzing hepatic enzymes and histopathological examination (in vivo) or lactate dehydrogenase (LDH) assay and morphological examination (in vitro). Global gene expression was assessed using microarray. In high dose TAA-treated group, there was centrilobular necrosis (in vivo) and cellular toxicity with an elevation of LDH (in vitro) at 24 h. Statistical analysis of global gene expression identified that there were similar numbers of altered genes found between in vivo and in vitro at each time points. Pathway analysis identified several common pathways existed between in vivo and in vitro system such as glutathione metabolism, bile acid biosynthesis, nitrogen metabolism, butanoate metabolism for hepatotoxicty caused by TAA. Our results suggest it may be feasible to develop toxicogenomics biomarkers by comparing in vivo and in vitro genomic profiles specific to TAA for application to prediction of liver toxicity.

• Horticultural Science & Technology
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• v.32 no.1
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• pp.91-99
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• 2014

The objective of this study is to identify cold-tolerance genes in Brassica rapa. In order to acheive this goal, we analyzed a KBGP-24K oligo chip data [BrEMD (B. rapa EST and Microarray Database)] using B. rapa ssp. pekinensis inbred line 'Chiifu' under cold stress condition ($4^{\circ}C$). Among 23,929 unigenes of B. rapa, 417 genes (1.7%) were primarily identified as cold responsive genes that were expressed over 5-fold higher than those of wild type control, and then a gene which has unknown function and has full length sequence was selected. It was named BrCSR (B. rapa Cold Stress Resistance). BrCSR was transformed using expression vector pSL101 to confirm whether BrCSR can enhance cold tolerance in tobacco plants. $T_1$ transgenic tobacco plants expressing BrCSR were selected by PCR and Southern hybridization analyses, and the function of BrCSR was characterized by expression level analysis and phenotype observation under cold stress condition. The expression level of BrCSR in transgenic tobacco plants increased up to about two folds in quantitative real-time RT-PCR assay and this was very similar to Northern blot hybridization analysis. Analysis of phenotypic characteristics clearly elucidated that transgenic tobaccos expressing BrCSR were more cold tolerant than wild type control under $4^{\circ}C$ treatment. Based on these results, we conclude that the over-expression of BrCSR might be closely related to the enhancement of cold tolerance.

• Journal of Life Science
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• v.19 no.9
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• pp.1294-1298
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• 2009

To investigate whether isoflavone genistein and daidzein could affect cancer cell viability, human colorectal HCT116 cells were incubated with genistein or daidzein in a dose-dependent manner. Genistein decreased cancer cell viability in a dose-dependent manner, whereas daidzein did not show dramatic cytotoxic effects. We also found that 71 genes were up-regulated more than 2-fold, whereas 64 genes were down-regulated more than 2-fold with 24 hr of $50{\mu}M$ genistein treatment by our previous microarray data. Among the up-regulated genes, we selected 3 genes (DKK1, ATF3 and NAG-1) and performed RT-PCR to confirm microarray data. The results of RT-PCR were highly correlated with those of the microarray experiment. In addition, we investigated whether a combination treatment of genistein and daidzein could affect cancer cell viability. Surprisingly, the combination treatment did show synergistic cytotoxic effects detected by MTS assay. The results of RT-PCR and real-time PCR indicate that a combination of genistein and daidzein can synergistically induce NAG-1 expression in HCT116 cells. This result implies that NAG-1 induction is highly associated with synergistic cytotoxic effects induced by a combination treatment of genistein and daidzein. Overall, these results may provide a clue in explaining the anti-cancer activity of soy bean in human colorectal cancer.

• Journal of Acupuncture Research
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• v.19 no.6
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• pp.111-124
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• 2002

A line of study reported that electroacupuncture(EA) modulate natural killer cell(NK Cell) activities. One report suggested that EA enhanced splenic interferon-gamma($IFN-{\gamma}$), interleukin-2(IL-2), and NK cell activity in Sprague-Dawley rats. Another study suggested that $IFN-{\gamma}$ mediates the up-regulation of NK cell activity, and endogenous ${\beta}$-endorphin secretion also play a role in the up-regulation of NK cell activity induced by EA stimulation. In order to better understand the molecular regulation underlying the activation of NK cell induced by EA, we have utilized cDNA microarray to elucidate how EA alters program of gene expression of spleen in rats. First, we divided three groups, group I was EA group treated with EA in restriction holder, group II was sham group with only holder stress, and last group III was control group with no treatment. We measured NK cell activity after EA stimulation three times for 2 days using $^{51}Cr$ release assay. Second, Biotin-labeled cDNA probes synthesized from EA group and sham group, were competitively hybridized to the microarray that contained variable genes. Such high-throughput screening has identified a number of EA-responsive gene candidates. Of these, we found that EA induced a subset of genes of genes that functionally could modulatory effects on NK cell activity. Genes(vascular cell adhesion molecule-1, protein-tyrosine kinase, CD94 mRNA) related to boost NK cell activity, were increased by EA And, genes(protein-tyrosine-phospatase mRNA, protein-tyrosine phosphatase(SHP-1) mRNA) related to inhibit NK cell activity, were decreased by EA. These EA-responsive genes may provide key insights from which to understand mechanisms of activation of NK cell induced by EA.

• Korean Journal of Head & Neck Oncology
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• v.33 no.1
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• pp.21-29
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• 2017

Methylation of CpG islands in the promoter region of genes acts as a significant mechanism of epigenetic gene silencing in head and neck squamous cell carcinoma (HNSCC). DNA methylation markers are particularly advantageous because DNA methylation is an early event in tumorigenesis, and the epigenetic modification, 5-methylcytosine, is a stable mark. In the present study, we assessed the genome-wide preliminary screening and were to identify novel methylation biomarker candidate in HNSCC. Genome-wide methylation analysis was performed on 10 HNSCC tumors using the Methylated DNA Isolation Assay (MeDIA) CpG island microarray. Validation was done using immunohistochemistry using tissue microarray of 135 independent HNSCC tumors. In addition, in vitro proliferation, migration/invasion assays, RT-PCR and immunoblotting were performed to elucidate molecular regulating mechanisms. Our preliminary validation using CpG microarray data set, immunohisto-chemistry for HNSCC tumor tissues and in vitro functional assays revealed that methylation of the Homeobox B5 (HOXB5) and H6 Family Homeobox 2 (HMX2) could be possible novel methylation biomarkers in HNSCC.

• Korean Journal of Microbiology
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• v.44 no.3
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• pp.169-177
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• 2008

Global gene expression of Deinococcus radiodurans, a highly radiation resistant bacterium, in response to excess copper was analyzed by using oligonucleotide microarray chip. Among 3,187 open reading frames of D. radiodurans, seventy genes showed a statistically significant expression ratio of at least 2-fold changes under growth conditions of excess copper; 64 genes were induced and 6 genes were reduced. Especially, two operons ($DRB0014{\sim}DRB0017$ and $DRB0125{\sim}DRB0121$) presumably involved in the iron transport and utilization were the most highly induced genes by excess copper. A quantitative real-time PCR assay revealed that DRB00l4 and DRB0125 are highly transcribed responding to excess copper and 2,2'-dipyridyl, an iron chelator. In addition, the transcription of both genes was not changed by excess iron and bathocuproine disulphonate, a copper chelator. These results suggested that the copper metabolism may be closely connected with the iron transport and utilization in D. radiodurans. However, the disruption of each gene, DRB00l4 and DRB0125, did not affect the copper and radiation resistance, the most well-known character of this organism.

• Korean Journal of Biological Psychiatry
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• v.12 no.1
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• pp.42-61
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• 2005

Objectives:The ginsenoside Rg1 and Rb1, the major components of ginseng saponin, have neurotrophic and neuroprotective effects including promotion of neuronal survival and proliferation, facilitation of learning and memory, and protection from ischemic injury and apoptosis. In this study, to investigate the molecular basis of the effects of ginsenoside on neuron, we analyzed gene expression profiling of SH-SY5Y human neuroblastoma cells treated with ginsenoside Rg1 or Rb1. Methods:SH-SY5Y cells were cultured and treated in triplicate with ginsenoside Rg1 or Rb1($80{\mu}M$, $40{\mu}M$, $20{\mu}M$). The proliferation rates of SH-SY5Y cells were determined by MTT assay and microscopic examination. We used a high density cDNA microarray chip that contained 8K human genes to analyze the gene expression profiles in SH-SY5Y cells. We analyzed using the Significance Analysis of Microarray(SAM) method for identifying genes on a microarray with statistically significant changes in expression. Results:Treatment of SH-SY5Y cells with $80{\mu}M$ ginsenoside Rg1 or Rb1 for 36h showed maximal proliferation compared with other concentrations or control. The results of the microarray experiment yielded 96 genes were upregulated(${\geq}$3 fold) in Rg1 treated cells and 40 genes were up-regulated(${\geq}$2 fold) in Rb1 treated cells. Treatment with ginsenoside Rg1 for 36h induced the expression of some genes associated with protein biosynthesis, regulation of transcription or translation, cell proliferation and growth, neurogenesis and differentiation, regulation of cell cycle, energy transport and others. Genes associated with neurogenesis and neuronal differentiation such as SCG10 and MLP increased in ginsenoside Rg1 treated cells, but such changes did not occur in Rb1-group. Conclusion:Our data provide novel insights into the gene mechanisms involved in possible role for ginsenoside Rg1 or Rb1 in mediating neuronal proliferation or cell viability, which can elicit distinct patterns of gene expression in neuronal cell line. Ginsenoside Rg1 have more broad and strong effects than ginsenoside Rb1 in gene expression and related cellular physiology. In addition, we suggest that SCG10 gene, which is known to be expressed in neuronal differentiation during development and neuronal regeneration during adulthood, may have a role in enhancement of activity dependent synaptic plasticity or cytoskeletal regulation following treatment of ginsenoside Rg1. Further, ginsenoside Rg1 may have a possible role in regeneration of injured neuron, promotion of memory, and prevention from aging or neuronal degeneration.

• Genomics & Informatics
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• v.11 no.4
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• pp.245-253
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• 2013

A radioresistant cell line was established by fractionated ionizing radiation (IR) and assessed by a clonogenic assay, flow cytometry, and Western blot analysis, as well as zymography and a wound healing assay. Microarray was performed to profile global expression and to search for differentially expressed genes (DEGs) in response to IR. H460R cells demonstrated increased cell scattering and acidic vesicular organelles compared with parental cells. Concomitantly, H460R cells showed characteristics of increased migration and matrix metalloproteinase activity. In addition, H460R cells were resistant to IR, exhibiting reduced expression levels of ionizing responsive proteins (p-p53 and ${\gamma}$-H2AX); apoptosis-related molecules, such as cleaved poly(ADP ribose) polymerase; and endoplasmic reticulum stress-related molecules, such as glucose-regulated protein (GRP78) and C/EBP-homologous protein compared with parental cells, whereas the expression of anti-apoptotic X-linked inhibitor of apoptosis protein was increased. Among DEGs, syntrophin beta 2 (SNTB2) significantly increased in H460R cells in response to IR. Knockdown of SNTB2 by siRNA was more sensitive than the control after IR exposure in H460, H460R, and H1299 cells. Our study suggests that H460R cells have differential properties, including cell morphology, potential for metastasis, and resistance to IR, compared with parental cells. In addition, SNTB2 may play an important role in radioresistance. H460R cells could be helpful in in vitro systems for elucidating the molecular mechanisms of and discovering drugs to overcome radioresistance in lung cancer therapy.